Materials for magnetizing cells and magnetic manipulation

Abstract

A material comprising positively and negatively charged nanoparticles, wherein one of said nanoparticles contained a magnetically responsive element, are combined with a support molecule, which is a long natural or synthetic molecule or polymer to make a magnetic nanoparticle assembly. When the magnetic nanoparticle assembly is combined with cells, it will magnetize those cells. The magnetized cells can then be washed to remove the magnetic nanoparticle assembly and the magnetized cells manipulated in a magnetic field.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application claims priority to U.S. Provisional Application No. 61 / 245,846, which was filed on Sep. 25, 2009 and is incorporated by reference in its entirety.FEDERALLY SPONSORED RESEARCH STATEMENT[0002]Not applicable.REFERENCE TO MICROFICHE APPENDIX[0003]Not applicable.FIELD OF THE INVENTION[0004]The invention relates to the fields of nanotechnology, materials, biosynthesis, medicine, cellular biology, and tissue engineering. More particularly, the compositions and methods of the present disclosure relate to methods of magnetizing cells, and 3D cell culturing, cell manipulation, and cell patterning using magnetic fields.BACKGROUND OF THE INVENTION[0005]Manipulating cells, controlling their environment, and promoting conditions that mimic or illicit in vivo or natural cellular or tissue responses is an area of intense research. In the area of stem cells and regenerative medicine there is a particular need for methods and materials tha...

AI Technical Summary

Benefits of technology

[0018]Generally speaking, the invention is a new material that allows cells to uptake or adsorb magnetically responsive elements, and thus be levitatable in cell culture when a magnetic field is applied. The materials include positively and negatively charged nanoparticles, one of which must contain one or more magnetically responsive elements, such as iron oxide. These nanoparticles are further combined with a polymer, preferably a natural or cell derived polymer, or other long molecule that acts as a support (herein called a “support molecule”) for the charged nanoparticles and the cells, holding the nanoparticles in place for their uptake or adsorption by the cells. The inclusion of both positive and negative nanoparticles allows intimate admixing of the nanoparticles and drives the assembly of the three components, thus ensuring even distribution and good uptake. The support molecule intimately combines all three components with the cells in fibrous mat-like structure that allows the cells to take up the magnetically responsive element.

[0019]After a period of incubation, the material can be washed away, allowing the cells to be manipulated in a magnetic field. An alternative step is to optimize or tune the uptake of magnetic material by increasing the ratio between the number of cells and the amount of magnetic nanoparticle. If a large number of cells are present they will uptake most of the magnetic nanoparticles, and the step of washing way any leftover material may no be necessary, particularly if the remaining support molecules and / or nanoparticles are non-toxic and / or beneficial to the cell. This is particularly true where the support molecules comprise one or more extracellular matrix protein, glycoprotein or polysaccharide. The magnetic particles are eventually lost from the cells, leaving them in a completely natural state.

[0020]In addition to simple 3D culturing, the magnetic field can be used to manipulate cell shape, patterns and motion. For example, the use of a toroidal (washer shaped) magnet can promote the cells to assemble into a similar shape or a tilted field can make the 3D cell culture thicker on one side. We have also created firm dense sheets of cells, by placing a strong magnet at the bottom of a culture dish for a period of growth. Simply reversing the field, allows the sheet to then be levitated and we can then continue growing the sheet in a 3D culture. It is also possible to combine various shapes and continue 3D culturing and thus create more complex shapes in a 3D culture.

[0021]The magnetically responsive element can be any element or molecule that will respond to a magnetic field, e.g., rare earth magnets (e.g., samarium cobalt (SmCo) and neodymium iron boron (NdFeB)), ceramic magnet materials (e.g., strontium ferrite), the magnetic elements (e.g., iron, cobalt, and nickel and their alloys and oxides). Particularly preferred are paramagnetic materials that react to a magnetic field, but are not magnets themselves, as this allows for easier assembly of the materials.

[0035]Our extensive testing of the above described system has shown there are a great number of improvements now made available over the prior art methods. First, the self assembly manufacturing chemistry makes the method simple and reproducible, and no specialized equipment is required for the manufacture of the magnetic nanoparticle assembly, or for subsequent cell magnetizing, manipulation or 3D culturing. The only requirements were a magnetic field, pipettes, containers and a hot plate. Thus, the method is compatible with large scale and high-throughput.

[0038]Levitating and culturing cells in 3D by magnetic levitation does not require any specialized or costly equipment or methods (such as for agitating or maintaining buoyancy) beyond standard 2D cell culturing requirements, and shape control of magnetically levitated 3D cell culture can be achieved by varying the magnetic field shape. Finally, and perhaps most importantly, the invention promotes rapid cell-cell interaction (scale of seconds and minutes) with levitation of cells and assembly into 3D multicellular clusters within minutes, and that complex culture structures can be made by manipulating the magnetic field and / or by magnetically bringing different cell types into contact.

Problems solved by technology

Many of these processes are not elastic or reversible, therefore, cells cannot return to their native state.

This introduces an artificial substrate with which cells interact, rather than rapidly promoting cell-cell interactions, and although an improvement over 2D culturing, the scaffolding is likely to perturb the cells and remains in the finished product.

Further, cells can grow on or in the microcarriers, but cells cannot be levitated in a manner where all around cell-cell contact / interaction is possible.

Nationally, there is a significant level of complexity involved in the fabrication of the microcarriers of Felder, which includes laborious chemistry and the need for complex equipment.

Buoyancy control also seems to be relevant to facilitate levitation, and is controlled by the infusion of glass bubbles into the microcarriers, again contributing to complexity and difficulty.

However, the shear stress resulting from agitation is known to cause cell damage.

Furthermore, agitation impairs any magnetic field shape control of 3D cultures.

The coating remains with the cells during culture, thus introducing an unnatural element in the culture and probably perturbing the cells.

The inventors contemplate the use of a biodegradable coating that could eventually be eliminated, but none are disclosed, so it is not known if this approach would be successful.

Finally, this system is cumbersome and not suitable for scale-up and high-throughput applications.

However, although able to produce sheets of cells, the cells are still grown on the bottom of a plate, and thus this is not true 3D culturing by magnetic levitation.

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